Chromatin architecture is a key determinant in the regulation of gene expression, and this architecture is strongly influenced by post-translational modifications of histones (Marks, P. A.; et al. Curr Opin Oncol 2001, 13 (6), 477-483; Luger, K.; et al. Nature 1997, 389 (6648), 251-260). Histone protein tails contain lysine residues that interact with the negative charges on the DNA backbone. These lysine-containing tails, consisting of up to 40 amino acid residues, protrude through the DNA strand, and act as a site for post-translational modification of chromatin, allowing alteration of higher order nucleosome structure (Jenuwein, T.; Allis, C. D. Science 2001, 293 (5532), 1074-1080). Multiple post-translational modifications of histones can mediate epigenetic remodeling of chromatin, with acetylation being the best characterized process (Johnstone, R. W. Nat Rev Drug Discov 2002, 1 (4), 287-299). Transcriptional repression is associated with specific CpG island DNA methylation and recruitment of histone deacetylases (HDACs) to gene promoters that cooperate in the epigenetic silencing of specific genes (Herman, J. G.; Baylin, S. B. N Engl J Med 2003, 349 (21), 2042-2054; Robertson, K. D. Oncogene 2001, 20 (24), 3139-3155). Normal mammalian cells exhibit an exquisite level of control of chromatin architecture by maintaining a balance between histone acetyltransferase (HAT) and HDAC activity (Shogren-Knaak, M.; et al. Science 2006, 311 (5762), 844-847).
In cancer, CpG island DNA promoter hypermethylation in combination with other chromatin modifications, including decreased activating marks and increased repressive marks on histone proteins 3 and 4, have been associated with the silencing of tumor suppressor genes (Baylin, S. B.; Ohm, J. E. Nat Rev Cancer 2006, 6 (2), 107-116). The important role of promoter CpG island methylation and its relationship to covalent histone modifications has recently been reviewed (Jones, P. A.; Baylin, S. B. Cell 2007, 128 (4), 683-692). As was mentioned above, the N-terminal lysine tails of histones can undergo numerous posttranslational modifications, including phosphorylation, ubiquitination, acetylation and methylation (Johnstone, R. W. Nat Rev Drug Discov 2002, 1 (4), 287-299; Shi, Y.; et al. Cell 2004, 119 (7), 941-953; Whetstine, J. R.; et al. Cell 2006, 125 (3), 467-481). To date, 17 lysine residues and 7 arginine residues on histone proteins have been shown to undergo methylation, and lysine methylation on histones can signal transcriptional activation or repression, depending on the specific lysine residue involved (Bannister, A. J.; et al. Nature 2005, 436 (7054), 1103-1106; Kouzarides, T. Curr Opin Genet Dev 2002, 12 (2), 198-209; Martin, C.; et al. Nat Rev Mol Cell Biol 2005, 6 (11), 838-849; Zhang, Y.; Reinberg, D. Genes Dev 2001, 15 (18), 2343-2360). All known histone lysine methyltransferases contain a conserved SET methyltransferase domain, and it has been shown that aberrant methylation of histones due to SET domain deregulation is linked to carcinogenesis (Schneider, R.; et al. Trends Biochem Sci 2002, 27 (8), 396-402). Histone methylation, once thought to be an irreversible process, has recently been shown to be a dynamic process regulated by the addition of methyl groups by histone methyltransferases and removal of methyl groups from mono- and dimethyllysines by lysine specific demethylase 1 (LSD1), and from mono-, di, and trimethyllysines by specific Jumonji C (JmjC) domain-containing demethylases (Shi, Y.; et al. Cell 2004, 119 (7), 941-953; Whetstine, J. R.; et al. Cell 2006, 125 (3), 467-481; Tsukada, Y.; Zhang, Y. Methods 2006, 40 (4), 318-326; Huarte, M.; et al. J Biol Chem 2007). Additional demethylases in the JmjC demethylase class are continuing to be identified (Liang, G.; et al. Nat Struct Mol Biol 2007, 14 (3), 243-245; Secombe, J.; et al. Genes Dev 2007, 21 (5), 537-551. Recent evidence suggests that LSD1 is required for maintenance of global DNA methylation, indicating that the LSD1-mediated demethylation is a general mechanism for transcriptional control (Wang, J.; et al. Nat Genet. 2009, 41 (1), 125-129).
A key positive chromatin mark found associated with promoters of active genes is histone 3 dimethyllysine 4 (H3K4me2) (Liang, G., et al. Proc Natl Acad Sci USA 2004, 101 (19), 7357-7362; Schneider, R.; et al. Nat Cell Biol 2004, 6 (1), 73-77). LSD1, also known as BHC110 and KDM1, catalyzes the oxidative demethylation of histone 3 methyllysine 4 (H3K4me1) and H3K4me2, and is associated with transcriptional repression. H3K4me2 is a transcription-activating chromatin mark at gene promoters, and demethylation of this mark by LSD1 may prevent expression of tumor suppressor genes important in human cancer (Huang, Y.; et al. Proc Natl Acad Sci USA 2007, 104 (19), 8023-8028). Thus, LSD1 is emerging as an important new target for the development of specific inhibitors as a new class of antitumor drugs (Stavropoulos, P.; Hoelz, A. Expert Opin Ther Targets 2007, 11 (6), 809-820).
To date, only a few existing compounds have been shown to act as inhibitors of LSD1. The active site structure of LSD1 has considerable sequence homology to monoamine oxidases A and B (MAO A and B), and to N1-acetylpolyamine oxidase (APAO) and spermine oxidase (SMO) (Shi, Y.; et al. Cell 2004, 119 (7), 941-953; Lee, M. G.; et al. Chem Biol 2006, 13 (6), 563-567; Schmidt, D. M.; McCafferty, D. G. Biochemistry 2007, 46 (14), 4408-4416). It has been shown that classical MAO inhibitors phenelzine and tranylcypromine inactivate nucleosomal demethylation by the recombinant LSD1/CoRest complex, and increase global levels of H3K4me2 in the P19 cell line. The synthetic substrate analogue aziridinyl-K4H31-21 reversibly inhibited LSD1 with an IC50 of 15.6 μM, while propargyl-K4H31-21 produced time-dependent inactivation with a Ki of 16.6 μM (Culhane, J. C.; et al. J Am Chem Soc 2006, 128 (14), 4536-4537). Propargyl-K4H31-21 was later shown to inactivate LSD1 through formation of a covalent adduct with the enzyme-bound flavin cofactor (Schmidt, D. M.; McCafferty, D. G. Biochemistry 2007, 46 (14), 4408-4416; Szewczuk, L. M.; et al. Biochemistry 2007, 46, 6892-6902). McCafferty et al. recently described the synthesis of a series of trans-2-arylcyclopropylamine analogues that inhibit LSD1 with K, values between 188 and 566 μM (Gooden, D. M.; et al. Bioorg Med Chem Lett 2008, 18 (10), 3047-3051). However, in all but one instance, these analogues were 1-2 orders of magnitude more potent against MAO A and MOA B. Most recently, Ueda and coworkers identified small molecule tranylcypromine derivatives that are selective for LSD1 over MAO-A and MAO-B, and Binda et al. described similar tranylcypromine analogues that exhibited partial selectivity between LSD1 and the newly identified histone demethylase LSD2 (Ueda, R.; et al. J Am Chem Soc 2009, 131 (48), 17536-17537; Binda, C.; et al. J Am Chem Soc 2010, 132, ePub 10.1021/ja101557k).
LSD1 was identified in part because its C-terminal domain shares significant sequence homology with the amine oxidases acetylpolyamine oxidase (APAO) and spermine oxidase (SMO) (Wang, Y.; et al. Biochem Biophys Res Commun 2003, 304 (4), 605-611). Several groups have identified amines, guanidines or similar analogues that act as selective modulators of these 2 amine oxidases (Wang, Y.; et al. Biochem Biophys Res Commun 2003, 304 (4), 605-611; Ferioli, M. E.; et al. Toxicol Appl Pharmacol 2004, 201 (2), 105-111; Casara, P.; et al. Tet. Letters 1984, 25, 1891-1894; Bellelli, A.; et al. Biochem Biophys Res Commun 2004, 322 (1), 1-8; Wang, Y.; et al. Cancer Chemother Pharmacol 2005, 56 (1), 83-90; Cona, A.; et al. Biochemistry 2004, 43 (12), 3426-3435; Stranska, J.; et al. Biochimie 2007, 89 (1), 135-144). We previously reported the synthesis of a novel series of (bis)guanidines and (bis)biguanides that are potent antitrypanosomal agents in vitro, with IC50 values against Trypanosoma brucei as low as 90 nM (Bi, X.; et al. Bioorg Med Chem Lett 2006, 16 (12), 3229-3232).
Because of the promising cellular effects of various (bis)guanidines and (bis)biguanides the synthesis and evaluation of additional analogues was undertaken.